The specific aims will be: (a) determination of the standard molal electrode potentials (E-o-m) for Ag-AgCl and AG-AgBr electrodes in 20, 30, 40 and 50 mass % glycerol-H2O and ethanol-H2O, at 25, 0, -5, -10, -15 and -20 degrees C; (b) calculation of standard pH*(s) values for reference buffers, such as Tricine Bicine, Bes, Mops, and Hepes (in isotonic saline and in modified Krebs solution) in the same mixed solvents at the same temperatures as (a); (c) establishment of an operational definition of pH* using emf cells with liquid junction (consisting of hydrogen and calomel electrodes) and the pH*(s) values from similar cells without liquid junction; (d) calibration of conventional glass-calomel pH cells for use in making meaningful and reliable pH measures in mixed solvents at subzero temperatures. The methodology involves three basic steps: (1) The first step is the measurement of the emf of the cell with the reference buffer solution in mixed solvent media, both with and without liquid junction. (2) The second step is the determination of PSH, which is the limit approached by PSH for each buffer solution as the halide molality (mx) is extrapolated to zero. (3) The final step is to calculate the standard pH values by using the expression pH(s) = PsHo+log gamma x, in which the activity coefficient of the halide ion (gamma x) is obtained from a conventional definition, as was done previously in a prior NIH grant. The long-term objectives are: (i) to originate a practical means for storage of tissues in unfrozen media with 0.16 m NaCl or modified Krebs solution at subzero temperatures; (ii) to provide a list of cryoprotective nonelectrolytes (such as ethanol, glycerol etc.) which are relatively nontoxic and effective in preserving tissues without injury; (iii) to eliminate the error due to liquid junction and to provide standard pH data for buffers in water and in mixed solvents, with and without liquid junctions; (iv) to establish a "universal" pH scale based on Eom and pK values.